Mouse models susceptible to HCoV-229E and HCoV-NL63 and cross protection from challenge with SARS-CoV-2.

Human coronavirus 229E (HCoV-229E) and NL63 (HCoV-NL63) are endemic causes of upper respiratory infections such as the "common cold" but may occasionally cause severe lower respiratory tract disease in the elderly and immunocompromised patients. There are no approved antiviral drugs or vaccines for these common cold coronaviruses (CCCoV). The recent emergence of COVID-19 and the possible cross-reactive antibody and T cell responses between these CCCoV and SARS-CoV-2 emphasize the need to develop experimental animal models for CCCoV. Mice are an ideal experimental animal model for such studies, but are resistant to HCoV-229E and HCoV-NL63 infections. Here, we generated 229E and NL63 mouse models by exogenous delivery of their receptors, human hAPN and hACE2 using replication-deficient adenoviruses (Ad5-hAPN and Ad5-hACE2), respectively. Ad5-hAPN- and Ad5-hACE2-sensitized IFNAR-/- and STAT1-/- mice developed pneumonia characterized by inflammatory cell infiltration with virus clearance occurring 7 d post infection. Ad5-hAPN- and Ad5-hACE2-sensitized mice generated virus-specific T cells and neutralizing antibodies after 229E or NL63 infection, respectively. Remdesivir and a vaccine candidate targeting spike protein of 229E and NL63 accelerated viral clearance of virus in these mice. 229E- and NL63-infected mice were partially protected from SARS-CoV-2 infection, likely mediated by cross-reactive T cell responses. Ad5-hAPN- and Ad5-hACE2-transduced mice are useful for studying pathogenesis and immune responses induced by HCoV-229E and HCoV-NL63 infections and for validation of broadly protective vaccines, antibodies, and therapeutics against human respiratory coronaviruses including SARS-CoV-2.

Cadherin-5 facilitated the differentiation of human induced pluripotent stem cells into sinoatrial node-like pacemaker cells by regulating ß-catenin.

Our study was conducted to investigate whether cadherin-5 (CDH5), a vascular endothelial cell adhesion glycoprotein, could facilitate the differentiation of human induced pluripotent stem cells (hiPSCs) into sinoatrial node-like pacemaker cells (SANLPCs), following previous findings of silk-fibroin hydrogel-induced direct conversion of quiescent cardiomyocytes into pacemaker cells in rats through the activation of CDH5. In this study, the differentiating hiPSCs were treated with CDH5 (40 ng/mL) between Day 5 and 7 during cardiomyocytes differentiation. The findings in the present study demonstrated that CDH5 stimulated the expression of pacemaker-specific markers while suppressing markers associated with working cardiomyocytes, resulting in an increased proportion of SANLPCs among hiPSCs-derived cardiomyocytes (hiPSC-CMs) population. Moreover, CDH5 induced typical electrophysiological characteristics resembling cardiac pacemaker cells in hiPSC-CMs. Further mechanistic investigations revealed that the enriched differentiation of hiPSCs into SANLPCs induced by CDH5 was partially reversed by iCRT14, an inhibitor of ß-catenin. Therefore, based on the aforementioned findings, it could be inferred that the regulation of ß-catenin by CDH5 played a crucial role in promoting the enriched differentiation of hiPSCs into SANLPCs, which presents a novel avenue for the construction of biological pacemakers in forthcoming research.

DNA methylase 1 influences temperature responses and development in the invasive pest Tuta absoluta.

DNA methylase 1 (Dnmt1) is an important regulatory factor associated with biochemical signals required for insect development. It responds to changes in the environment and triggers phenotypic plasticity. Meanwhile, Tuta absoluta Meyrick (Lepidoptera: Gelechiidae)-a destructive invasive pest-can rapidly invade and adapt to different habitats; however, the role of Dnmt1 in this organism has not been elucidated. Accordingly, this study investigates the mechanism(s) underlying the rapid adaptation of Tuta absoluta to temperature stress. Potential regulatory genes were screened via RNAi (RNA interference), and the DNA methylase in Tuta absoluta was cloned by RACE (Rapid amplification of cDNA ends). TaDnmt1 was identified as a potential regulatory gene via bioinformatics; its expression was evaluated in response to temperature stress and during different development stages using real-time polymerase chain reaction. Results revealed that TaDnmt1 participates in hot/cold tolerance, temperature preference and larval development. The full-length cDNA sequence of TaDnmt1 is 3765 bp and encodes a 1254 kDa protein with typical Dnmt1 node-conserved structural features and six conserved DNA-binding active motifs. Moreover, TaDnmt1 expression is significantly altered by temperature stress treatments and within different development stages. Hence, TaDnmt1 likely contributes to temperature responses and organismal development. Furthermore, after treating with double-stranded RNA and exposing Tuta absoluta to 35°C heat shock or -12°C cold shock for 1 h, the survival rate significantly decreases; the preferred temperature is 2°C lower than that of the control group. In addition, the epidermal segments become enlarged and irregularly folded while the surface dries up. This results in a significant increase in larval mortality (57%) and a decrease in pupation (49.3%) and eclosion (50.9%) rates. Hence, TaDnmt1 contributes to temperature stress responses and temperature perception, as well as organismal growth and development, via DNA methylation regulation. These findings suggest that the rapid geographic expansion of T absoluta has been closely associated with TaDnmt1-mediated temperature tolerance. This study advances the research on 'thermos Dnmt' and provides a potential target for RNAi-driven regulation of Tuta absoluta.

A remote sensing method for mapping alpine grasslines based on graph-cut.

Climate change has induced substantial shifts in vegetation boundaries such as alpine treelines and shrublines, with widespread ecological and climatic influences. However, spatial and temporal changes in the upper elevational limit of alpine grasslands ("alpine grasslines") are still poorly understood due to lack of field observations and remote sensing estimates. In this study, taking the Tibetan Plateau as an example, we propose a novel method for automatically identifying alpine grasslines from multi-source remote sensing data and determining their positions at 30-m spatial resolution. We first identified 2895 mountains potentially having alpine grasslines. On each mountain, we identified a narrow area around the upper elevational limit of alpine grasslands where the alpine grassline was potentially located. Then, we used linear discriminant analysis to adaptively generate from Landsat reflectance features a synthetic feature that maximized the difference between vegetated and unvegetated pixels in each of these areas. After that, we designed a graph-cut algorithm to integrate the advantages of the Otsu and Canny approaches, which was used to determine the precise position of the alpine grassline from the synthetic feature image. Validation against alpine grasslines visually interpreted from a large number of high-spatial-resolution images showed a high level of accuracy (R2 , .99 and .98; mean absolute error, 22.6 and 36.2 m, vs. drone and PlanetScope images, respectively). Across the Tibetan Plateau, the alpine grassline elevation ranged from 4038 to 5380 m (5th-95th percentile), lower in the northeast and southeast and higher in the southwest. This study provides a method for remotely sensing alpine grasslines for the first-time at large scale and lays a foundation for investigating their responses to climate change.

Integrative analysis of circRNA networks in postoperative cognitive dysfunction.

OBJECTIVE: In the quest to decipher the molecular intricacies of Postoperative Cognitive Dysfunction (POCD), this study focused on circular RNA (circRNA) and their regulatory networks. MATERIALS AND METHODS: Analyzing the Gene Expression Omnibus Series (GSE) 147277 dataset, we pinpointed 10 differentially expressed circRNAs linked to POCD. RESULTS: The ensuing competing endogenous RNA (ceRNA) network, featuring pivotal players like Homo sapiens(hsa)_circ_0003424 and hsa-miR-193b-5p, provided a comprehensive understanding of the molecular players at play in POCD. CONCLUSION: Additionally, the Protein-Protein Interaction (PPI) network spotlighted 10 core Hub genes, including phosphatase and tensin homolog (PTEN) and signal transducer and activator of transcription 3(STAT3), shedding light on potential therapeutic targets.

Inhibition of intermittent calcium-activated potassium channel (SK4) attenuates Ang II-induced hypertrophy of human-induced stem cell-derived cardiomyocytes via targeting Ras-Raf-MEK1/2-ERK1/2 and CN-NFAT signaling pathways.

Cardiac hypertrophy caused by angiotensin II (Ang II) is essential for the pathological process of heart failure. The intermediate calcium-activated potassium channel (SK4) has been shown to be involved in the process of the inflammatory response, cell proliferation, and apoptosis. However, the role of SK4 in cardiac hypertrophy has not been elucidated. Cardiac hypertrophy in human-induced pluripotent stem cells-derived cardiomyocytes (HiPSC-CMs) was induced by Ang II. Cells were transfected with SK4 adenovirus or treated with SK4 inhibitor (TRAM-34). TUNEL staining was used to assess the levels of apoptosis. Real-time polymerase chain reaction and Western blot analysis were used to measure messenger RNA (mRNA) and protein levels, respectively. The present results showed that SK4 expression was upregulated in HiPSC-CMs stimulated by Ang II. The downregulation of SK4 by a specific inhibitor TRAM-34 markedly ameliorated cardiac hypertrophy (reflected by the mRNA levels of atrial natriuretic peptide, brain natriuretic peptide, and ß-myosin heavy chain) and apoptosis (reflected by the level of Caspase 3, Bax, and Bcl-2) induced by Ang II treatment. The action of SK4 in cardiac hypertrophy was mediated by Ras-Raf-mitogen-activated protein kinases 1/2 (MEK1/2)-extracellular-regulated protein kinases 1/2 (ERK1/2) and calcineurin (CN)-nuclear factors of activated T cells (NFAT) activation. Our studies demonstrated that inhibition of SK4 significantly alleviated cardiac hypertrophy induced by Ang II in hiPSC-CMs by targeting Ras-Raf-MEK1/2-ERK1/2 signaling and CN-NFAT signaling pathway. Our studies suggest that SK4 may serve as a potential therapeutic target that could delay hypertrophy.

The paired-related homeobox protein 1 promotes cardiac fibrosis via the Twist1-Prrx1-tenascin-C loop.

Cardiac fibrosis is a common pathology in the advanced stage of cardiovascular diseases, which leads to cardiac systolic and diastolic dysfunction. It is important to prevent cardiac fibrosis during myocardial injury. The transcription factor Prrx1 is involved in cancer-associated fibrosis and other organ fibrosis. However, the role and mechanism of Prrx1 in cardiac fibrosis deserves further exploration. We identified that overexpressed Prrx1 promoted the proliferation and migration of cardiac fibroblasts, and transform cardiac fibroblasts to myofibroblasts in vitro. We demonstrated that the expression of Prrx1 is upregulated in TGF-ß1-treated fibroblasts. And silencing Prrx1 could attenuate cardiac fibrosis induced by TGF-ß1 in vitro. In addition, a Twist1-paired-related homeobox 1 (Prrx1)-tenascin-C (TNC) positive feedback loop (PFL) combined with Twist1, Prrx1, and TNC activated fibroblasts, which was the mechanism the Prrx1 in cardiac fibrosis. In conclusion, our findings showed that the deficiency of Prrx1 attenuated cardiac fibrosis in vitro and reveal a novel Twist1-Prrx1-TNC PFL in the regulation of cardiac fibrosis.

The method of sinus node-like pacemaker cells from human induced pluripotent stem cells by BMP and Wnt signaling.

The embryonic development of sinus nodes (SAN) is co-regulated by multiple signaling pathways. Among these, the bone morphogenetic protein (BMP) and Wnt signaling pathways are involved in the development of SAN. In this study, the effects of BMP and Wnt signaling on the differentiation of SAN-like pacemaker cells (SANLPCs) were investigated. Human induced pluripotent stem cells (hiPSCs) were divided into four groups: control, BMP4, CHIR-3, and BMP4 + CHIR (CHIR: a Wnt signaling activator). The samples were tested at day (D) 15 of differentiation. The final protocol for the activation of BMP signaling at D0-D3 and reactivation of Wnt signaling at D5-D7 in the differentiation of hiPSCs were determined. The results showed that the mRNA levels of pacemaker markers (TBX18, SHOX2, TBX3, HCN4, and HCN1) were higher in the BMP4 + CHIR group than in the control group, and working myocardial genes were downregulated. The immunofluorescence assay revealed that the expression of SHOX2 and HCN4 increased in the BMP4 + CHIR group compared to that in the other groups. In addition, the results of patch clamps revealed that a funny current of higher density and typical SAN action potentials were recorded, except in the control group, in which the L-type calcium current was higher in the BMP4 + CHIR group than in the other groups. Finally, the proportion of SANLPCs (cTnT+ NKX2.5-) was further enhanced by the combination of BMP4 and CHIR treatment. In summary, the combination of BMP and Wnt signaling promotes the differentiation of SANLPCs from hiPSCs.

Electrocatalytic Deep Dehalogenation and Mineralization of Florfenicol: Synergy of Atomic Hydrogen Reduction and Hydroxyl Radical Oxidation over Bifunctional Cathode Catalyst.

It is difficult to achieve deep dehalogenation or mineralization for halogenated antibiotics using traditional reduction or oxidation processes, posing the risk of microbial activity inhibition and bacterial resistance. Herein, an efficient electrocatalytic process coupling atomic hydrogen (H*) reduction with hydroxyl radical (•OH) oxidation on a bifunctional cathode catalyst is developed for the deep dehalogenation and mineralization of florfenicol (FLO). Atomically dispersed NiFe bimetallic catalyst on nitrogen-doped carbon as a bifunctional cathode catalyst can simultaneously generate H* and •OH through H2O/H+ reduction and O2 reduction, respectively. The H* performs nucleophilic hydro-dehalogenation, and the •OH performs electrophilic oxidization of the carbon skeleton. The experimental results and theoretical calculations indicate that reductive dehalogenation and oxidative mineralization processes can promote each other mutually, showing an effect of 1 + 1 > 2. 100% removal, 100% dechlorination, 70.8% defluorination, and 65.1% total organic carbon removal for FLO are achieved within 20 min (C0 = 20 mg·L-1, -0.5 V vs SCE, pH 7). The relative abundance of the FLO resistance gene can be significantly reduced in the subsequent biodegradation system. This study demonstrates that the synergy of reduction dehalogenation and oxidation degradation can achieve the deep removal of refractory halogenated organic contaminants.

Precipitation and local adaptation drive spatiotemporal variations of aboveground biomass and species richness in Tibetan alpine grasslands.

The Tibetan Plateau contains the highest and largest alpine pasture in the world, which is adapted to the cold and arid climate. It is challenging to understand how the vast alpine grasslands respond to climate change. We aim to test the hypothesis that there is local adaptation in elevational populations of major plant species in Tibetan alpine grasslands, and that the spatiotemporal variations of aboveground biomass (AGB) and species richness (S) can be mainly explained by climate change only when the effect of local adaptation is removed. A 7-year reciprocal transplant experiment was conducted among the distribution center (4950 m), upper (5200 m) and lower (4650 m) limits of alpine Kobresia meadow in central Tibetan Plateau. We observed interannual variations in S and AGB of 5 functional groups and 4 major species, and meteorological factors in each of the three elevations during 2012-2018. Relationships between interannual changes of AGB and climatic factors varied greatly with elevational populations within a species. Elevation of population origin generally had a greater or an equal contribution to interannual variation in AGB of the 4 major species, compared to temperature and precipitation effects. While the effect of local adaptation was removed by calculating differences in AGB and S between elevations of migration and origin, relative changes in AGB and S were mainly explained by precipitation change rather than by temperature change. Our data support the hypothesis, and further provide evidence that the monsoon-adapted alpine grasslands are more sensitive to precipitation change than to warming.

Muscone attenuates susceptibility to ventricular arrhythmia by inhibiting NLRP3 inflammasome activation in rats after myocardial infarction.

Fibrosis and abnormal expression of connexin 43 (Cx43) in the ventricle play vital roles in ventricular arrhythmias (VAs) after myocardial infarction (MI). Muscone, an active monomer of heart-protecting musk pill, has various biological activities, but its effect on susceptibility to VAs in rats with MI has not been determined. In the present study, we investigated the effects of muscone on ventricular inflammation, fibrosis, Cx43 expression, and the occurrence of VAs after MI. An MI model was established by ligating the proximal left anterior descending coronary artery. Then, the MI model rats were administered muscone (2 mg/kg/day) or vehicle (saline)via intragastric injection for 14 days. Cardiac function was evaluated by echocardiography, and an in vivo electrophysiological study was performed on Day 14. Cardiac inflammation, fibrosis, and Cx43 expression were determined by histochemical analysis and western blot analysis. Our results indicated that muscone treatment significantly improved cardiac function and inhibited ventricular inflammation, fibrosis, and nucleotide-binding oligomerization domain, leucine-rich repeat, and pyrin domain containing 3 (NLRP3) inflammasome activation. Electrocardiogrphy and electrophysiology studies showed that muscone shortened the QRS interval, QT interval, QTc interval, and action potential duration; prolonged the effective refractory period; and reduced susceptibility to VAs in rats after MI. Furthermore, Cx43 expression in the BZ was increased by muscone treatment, and this change was coupled by inhibition of the NLRP3/IL-1ß/p38 MAPK pathway. Taken together, our results demonstrated that muscone reduces susceptibility to VA, mainly by decreasing ventricular inflammation and fibrosis, and attenuates abnormal Cx43 expression by inhibiting NLRP3 inflammasome activation after myocardial infarction in rats.

Research on a Precision Calibration Model of a Flexible Strain Sensor Based on a Variable Section Cantilever Beam.

The flexible strain sensor's measuring range is usually over 5000 µÎµ, while the conventional variable section cantilever calibration model has a measuring range within 1000 µÎµ. In order to satisfy the calibration requirements of flexible strain sensors, a new measurement model was proposed to solve the inaccurate calculation problem of the theoretical strain value when the linear model of a variable section cantilever beam was applied to a large range. The nonlinear relationship between deflection and strain was established. The finite element analysis of a variable section cantilever beam with ANSYS shows that the linear model's relative deviation is as high as 6% at 5000 µÎµ, while the relative deviation of the nonlinear model is only 0.2%. The relative expansion uncertainty of the flexible resistance strain sensor is 0.365% (k = 2). Simulation and experimental results show that this method solves the imprecision of the theoretical model effectively and realizes the accurate calibration of a large range of strain sensors. The research results enrich the measurement models and calibration models for flexible strain sensors and contribute to the development of strain metering.

Calibration Model Optimization for Strain Metrology of Equal Strength Beams Using Deflection Measurements.

Strain sensors, especially fiber Bragg grating (FBG) sensors, are of great importance in structural health monitoring, mechanical property analysis, and so on. Their metrological accuracy is typically evaluated by equal strength beams. The traditional strain calibration model using the equal strength beams was built based on an approximation method by small deformation theory. However, its measurement accuracy would be decreased while the beams are under the large deformation condition or under high temperature environments. For this reason, an optimized strain calibration model is developed for equal strength beams based on the deflection method. By combining the structural parameters of a specific equal strength beam and finite element analysis method, a correction coefficient is introduced into the traditional model, and an accurate application-oriented optimization formula is obtained for specific projects. The determination method of optimal deflection measurement position is also presented to further improve the strain calibration accuracy by error analysis of the deflection measurement system. Strain calibration experiments of the equal strength beam were carried out, and the error introduced by the calibration device can be reduced from 10 µÎµ to less than 1 µÎµ. Experimental results show that the optimized strain calibration model and the optimum deflection measurement position can be employed successfully under large deformation conditions, and the deformation measurement accuracy is improved greatly. This study is helpful to effectively establish metrological traceability for strain sensors and furthermore improve the measurement accuracy of strain sensors in practical engineering scenarious.

Dapagliflozin reduces the vulnerability of rats with pulmonary arterial hypertension-induced right heart failure to ventricular arrhythmia by restoring calcium handling.

BACKGROUND: Malignant ventricular arrhythmia (VA) is a major contributor to sudden cardiac death (SCD) in patients with pulmonary arterial hypertension (PAH)-induced right heart failure (RHF). Recently, dapagliflozin (DAPA), a sodium/glucose cotransporter-2 inhibitor (SGLT2i), has been found to exhibit cardioprotective effects in patients with left ventricular systolic dysfunction. In this study, we examined the effects of DAPA on VA vulnerability in a rat model of PAH-induced RHF. METHODS: Rats randomly received monocrotaline (MCT, 60 mg/kg) or vehicle via a single intraperitoneal injection. A day later, MCT-injected rats were randomly treated with placebo, low-dose DAPA (1 mg/kg/day), or high-dose (3 mg/kg/day) DAPA orally for 35 days. Echocardiographic analysis, haemodynamic experiments, and histological assessments were subsequently performed to confirm the presence of PAH-induced RHF. Right ventricle (RV) expression of calcium (Ca2+) handling proteins were detected via Western blotting. RV expression of connexin 43 (Cx43) was determined via immunohistochemical staining. An optical mapping study was performed to assess the electrophysiological characteristics in isolated hearts. Cellular Ca2+ imaging from RV cardiomyocytes (RVCMs) was recorded using Fura-2 AM or Fluo-4 AM. RESULTS: High-dose DAPA treatment attenuated RV structural remodelling, improved RV function, alleviated Cx43 remodelling, increased the conduction velocity, restored the expression of key Ca2+ handling proteins, increased the threshold for Ca2+ and action potential duration (APD) alternans, decreased susceptibility to spatially discordant APD alternans and spontaneous Ca2+ events, promoted cellular Ca2+ handling, and reduced VA vulnerability in PAH-induced RHF rats. Low-dose DAPA treatment also showed antiarrhythmic effects in hearts with PAH-induced RHF, although with a lower level of efficacy. CONCLUSION: DAPA administration reduced VA vulnerability in rats with PAH-induced RHF by improving RVCM Ca2+ handling.

Tbx18 promoted the conversion of human-induced pluripotent stem cell-derived cardiomyocytes into sinoatrial node-like pacemaker cells.

Sinoatrial node (SAN) pacemaker cells originate from T-box transcription factor 18 (Tbx18)-expressing progenitor cells. The present study aimed to investigate whether overexpression of human transcription factor Tbx18 could reprogram human-induced pluripotent stem cell-derived cardiomyocytes (hiPSC-CMs) into SAN-like pacemaker cells (SANLPCs) in vitro. In the study, hiPSCs were first differentiated into hiPSC-CMs through regulating the Wnt/ß-catenin pathway, then purified hiPSC-CMs were transfected by Tbx18 adenovirus (Tbx18-CMs group) or green fluorescent protein (GFP) adenovirus (GFP-CMs group). The beating frequency of the Tbx18-CMs group was significantly higher than that of the hiPSC-CMs group and GFP-CMs group. Compared with the other two groups, the expression levels of hyperpolarization-activated cyclic nucleotide-gated potassium channel isoform 4, connexin-45 in the Tbx18-CMs group were markedly upregulated, while the expressions of transcription factor NKX2.5, CX43 were significantly downregulated. Whole-cell patch-clamp results illustrated that action potential and "funny" current (If ) similar to SAN pacemaker cells could be recorded in the Tbx18-CMs group. In conclusion, this present study demonstrated that overexpression of Tbx18 promoted the conversion of hiPSC-CMs into SANLPCs.

DnaJ, a promising vaccine candidate against Ureaplasma urealyticum infection.

Ureaplasma urealyticum (U. urealyticum, Uu) is a common sexually transmitted pathogen that is responsible for diseases such as non-gonococcal urethritis, chorioamnionitis, and neonatal respiratory diseases. The rapid emergence of multidrug-resistant bacteria threatens the effective treatment of Uu infections. Considering this, vaccination could be an efficacious medical intervention to prevent Uu infection and disease. As a highly conserved molecular chaperone, DnaJ is expressed and upregulated by pathogens soon after infection. Here, we assessed the vaccine potential of recombinant Uu-DnaJ in a mouse model and dendritic cells. Results showed that intramuscular administration of DnaJ induced robust humoral- and T helper (Th) 1 cell-mediated immune responses and protected against genital tract infection, inflammation, and the pathologic sequelae after Uu infection. Importantly, the DnaJ protein also induced the maturation of mouse bone marrow-derived dendritic cells (BMDCs), ultimately promoting naïve T cell differentiation toward the Th1 phenotype. In addition, adoptive immunization of DnaJ-pulsed BMDCs elicited antigen-specific Immunoglobulin G2 (IgG2) antibodies as well as a Th1-biased cellular response in mice. These results support DnaJ as a promising vaccine candidate to control Uu infections. KEY POINTS: • A novel recombinant vaccine was constructed against U. urealyticum infection. • Antigen-specific humoral and cellular immune responses after DnaJ vaccination. • Dendritic cells are activated by Uu-DnaJ, which results in a Th1-biased immune response.

Enzymatic Enantioselective anti-Markovnikov Hydration of Aryl Alkenes.

The addition of water to alkenes is an important method for the synthesis of alcohols, but the regioselectivity of acid-catalyzed hydration of terminal alkenes yields secondary alcohols according to Markovnikov's rule, making it difficult to obtain primary alcohols. Here we report a styrene monooxygenase that catalyzes the anti-Markovnikov hydration of the terminal aryl alkenes under anaerobic conditions. This hydration provides primary alcohols in good yields (up to 100 %), excellent anti-Markovnikov regioselectivity (>99 : 1), and good enantiomeric purity (60-83 % ee). Residues Asn46, Asp100, and Asn309 are essential for catalysis suggesting an acid-base mechanism with a carbanion-like intermediate that could account for the anti-Markovnikov regioselectivity. Our work reveals a new enzymatic tool with unusual regioselectivity based on the promiscuous catalytic activity of a monooxygenase.

Long-term observation of catheter ablation vs. pharmacotherapy in the management of persistent and long-standing persistent atrial fibrillation (CAPA study).

AIMS: The roles of radiofrequency catheter ablation (RFCA) and pharmacotherapy in treating persistent and long-standing persistent atrial fibrillation (AF) have not been sufficiently investigated. We conducted a multicentre, randomized, controlled trial to compare the effects of RFCA and pharmacotherapy on the prognosis of these patients. METHODS AND RESULTS: A total of 648 patients with persistent and long-standing persistent AF were enrolled from 30 centres and randomized to either the ablation group (n = 327) or the pharmacotherapy group (n = 321). After 54.2 ± 10.6 months of follow-up, the primary endpoints occurred significantly more rarely in the ablation group than in the pharmacotherapy group (10.4% vs. 17.4%; hazard ratio 0.59, 95% confidence interval 0.48-0.75; P < 0.001). The incidence of stroke/transient ischaemic attack (TIA) was significantly lower in the ablation group (4.2% vs. 7.2%, P < 0.001). Likewise, the incidence of new-onset congestive heart failure (CHF) was lower in the ablation group (2.8% vs. 7.2%, P < 0.001). More patients had sinus rhythm in the ablation group than in the pharmacotherapy group (60.6% vs. 20.9%, P < 0.001), but fewer patients were on antiarrhythmic drugs (24.4% vs. 41.6%, P < 0.001) and warfarin (60.8% vs. 83.9%, P = 0.001). Both the 6-min walk distance and the quality of life (QoL) were improved in the ablation group at the end of follow-up. CONCLUSION: In patients with persistent and long-standing persistent AF, RFCA-based treatment was superior to pharmacotherapy in decreasing stroke/TIA and new-onset CHF and improving QoL.

Fluvastatin attenuated ischemia/reperfusion-induced autophagy and apoptosis in cardiomyocytes through down-regulation HMGB1/TLR4 signaling pathway.

Fluvastatin, a traditional fat-decreasing drug, is widely used for curing cardiovascular disease. Previous reports demonstrated that fluvastatin pretreatment protected against myocardial ischemia/reperfusion (I/R) by inhibiting TLR4 signaling pathway and/or reducing proinflammatory cytokines. However, whether fluvastatin has a cardioprotective effect against apoptosis and autophagy remains unknown. This study aims to evaluate whether the cardioprotective role of fluvastatin in I/R is mediated by high-mobility group box 1 (HMGB1)/toll-like receptor 4 (TLR4) pathway via anti-apoptotic and anti-autophagic functions. Sprague-Dawley rats were anesthetized, artificially ventilated and subjected to 30 min of coronary occlusion, followed by 4 h of reperfusion. The animals were randomized into four groups: (i) Sham operation; (ii) I/R; (iii) I/R + low-dosage fluvastatin (10 mg/kg); and (iv) I/R + high-dosage fluvastatin (20 mg/kg). After reperfusion, the hemodynamic parameters, myocardial infarct size, structural alteration of myocardium, apoptosis index, pro-inflammatory cytokine production, Beclin-1, Light chain 3 (LC3), HMGB1, TLR4 and Nuclear factor kappa B (NF-κB) protein levels were measured and recorded. It was found that fluvastatin preconditioning improved left ventricular dysfunction, reduced HMGB1/TLR4/NF-κB expressions, and inhibited cardiomyocyte apoptosis, autophagy, and inflammation reaction. Moreover, treatment with fluvastatin ameliorated myocardial injury by reducing infarct size, causing less damage to cardiac structure, downregulating autophagy-related protein expression and releasing pro-inflammation mediators. Our findings indicate that fluvastatin exerts beneficial effects on cardiac ischemic damage, which may be associated with its anti-autophagic and anti-apoptotic functions via inhibition of HMGB1/TLR4-related pathway during I/R injury.

Left Stellate Ganglion Ablation Inhibits Ventricular Arrhythmias through Macrophage Regulation in Canines with Acute Ischemic Stroke.

AIMS: To investigate the potential mechanism of ventricular arrhythmias (VAs) after acute ischemic stroke and explore the effects of left stellate gangling (LSG) ablation on VAs induced by stroke in canines. Materials and Methods: Twenty canines were randomly divided into the sham-operated group (n=6), AS group (n=7) and SGA group (n=7). Cerebral ischemic model was established in the AS group and the SGA group by right acute middle cerebral artery occlusion (MCAO). LSG ablation was performed in the SGA group as soon as MCAO. After 3 days, atrial electrophysiology and neural activity were measured in vivo. The levels of norepinephrine (NE) in plasma and ventricle were detected by ELISA. The levels of monocyte chemotactic protein-1 (MCP-1), tumor necrosis factor-α (TNF-α) and NF-κB p65 in ventricle were detected by western blotting. The pro-inflammatory polarization of macrophages in ventricle was detected by immunofluorescence. Results: Higher ventricular tachycardia (VT) inducibility and lower ventricular fibrillation threshold (VFT) were observed in the AS group compared with those in the sham-operated group, associated with higher LSG activity and NE levels, increased number of M1 macrophages and secretion of inflammatory cytokines in ventricle (all P<0.001). Compared with the AS group, the SGA group had lower VT inducibility and higher VFT, combined with lower NE levels, and reduced number of M1 macrophages and secretion of inflammatory cytokines in ventricle (all P<0.001). Conclusion: LSG ablation could reduce VAs vulnerability after acute stroke by preventing the macrophages polarization and activation induced by sympathetic hyperactivity.